DNA Damage, n-3 Long-Chain PUFA Levels and Proteomic Profile in Brazilian Children and Adolescents.

Fatty acids play a significant role in maintaining cellular and DNA protection and we previously found an inverse relationship between blood levels of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) and DNA damage. The aim of this study was to explore differences in proteomic profiles, for 117 pro-inflammatory proteins, in two previously defined groups of individuals with different DNA damage and EPA and DHA levels. Healthy children and adolescents (n = 140) aged 9 to 13 years old in an urban area of Brazil were divided by k-means cluster test into two clusters of DNA damage (tail intensity) using the comet assay (cluster 1 = 5.9% ± 1.2 and cluster 2 = 13.8% ± 3.1) in our previous study. The cluster with higher DNA damage and lower levels of DHA (6.2 ± 1.6 mg/dL; 5.4 ± 1.3 mg/dL, p = 0.003) and EPA (0.6 ± 0.2 mg/dL; 0.5 ± 0.1 mg/dL, p < 0.001) presented increased expression of the proteins CDK8-CCNC, PIK3CA-PIK3R1, KYNU, and PRKCB, which are involved in pro-inflammatory pathways. Our findings support the hypothesis that low levels of n-3 long-chain PUFA may have a less protective role against DNA damage through expression of pro-inflammatory proteins, such as CDK8-CCNC, PIK3CA-PIK3R1, KYNU, and PRKCB.

Enzymes that hydrolyze adenine nucleotides in platelets and polymorphisms in the alpha2 gene of integrin alpha2beta1 in patients with von Willebrand disease.

Von Willebrand disease (VWD) is one of the most common inherited bleeding diseases caused by a qualitative or quantitative deficiency of the von Willebrand factor (FvW). FvW is a multimeric glycoprotein synthesized by megakaryocytes and endothelial cells and it is present in the subendothelial matrix, blood plasma, platelets, and endothelium. This glycoprotein plays an important role in thrombus formation by initiating platelet adhesion to sites of injury as well as platelet aggregation. The aim of this study was to evaluate the activities of enzymes that hydrolyze adenine nucleotides in platelets, ristocetin-induced platelet aggregation (RIPA), and polymorphisms of the alpha2 gene of alpha2beta1 integrin from VWD patients. Platelet nucleoside triphosphate diphosphohydrolase (NTPDase), 5'-nucleotidase, and ecto-nucleotide pyrophosphatase/phosphodiesterase (E-NPP) activities were verified in 14 VWD patients. For RIPA determination, a final concentration of 1.25 mg/ml of ristocetin was used. Polymorphisms of the alpha2 gene were analyzed through PCR. Platelet NTPDase and E-NPP were decreased in VWD patients. 5'-Nucleotidase activity was not statistically significant between controls and VWD patients. RIPA was significantly reduced, with an allelic frequency of 78.57% for 807C in VWD patients. Our results indicated reduced platelet NTPDase and E-NPP activities which might be related to the low platelet adhesiveness. The prevalence of the 807C allele might account for the variability in bleeding in VWD.

Temporal variations of adenosine metabolism in human blood.

Eight diurnally active (06:00-23:00 h) subjects were adapted for 2 days to the room conditions where the experiments were performed. Blood sampling for adenosine metabolites and metabolizing enzymes was done hourly during the activity span and every 30 min during sleep. The results showed that adenosine and its catabolites (inosine, hypoxanthine, and uric acid), adenosine synthesizing (S-adenosylhomocysteine hydrolase and 5'-nucleotidase), degrading (adenosine deaminase) and nucleotide-forming (adenosine kinase) enzymes as well as adenine nucleotides (AMP, ADP, and ATP) undergo statistically significant fluctuations (ANOVA) during the 24 h. However, energy charge was invariable. Glucose and lactate chronograms were determined as metabolic indicators. The same data analyzed by the chi-square periodogram and Fourier series indicated ultradian oscillatory periods for all the metabolites and enzymatic activities determined, and 24-h oscillatory components for inosine, hypoxanthine, adenine nucleotides, glucose, and the activities of SAH-hydrolase, 5'-nucleotidase, and adenosine kinase. The single cosinor method showed significant oscillatory components exclusively for lactate. As a whole, these results suggest that adenosine metabolism may play a role as a biological oscillator coordinating and/or modulating the energy homeostasis and physiological status of erythrocytes in vivo and could be an important factor in the distribution of purine rings for the rest of the organism.

Estudo das proteases do glóbulo vermelho humano de indivíduos normais e de portadores de hipertensäo / Study of the proteases of the human erythrocyte in normal patients and in hypertensive patients

Foram estudados indivíduos normais e portadores de hipertensäo essencial com a finalidade de verificar a presenca de proteases ligadas à membrana e também presentes no citosol do glóbulo vermelho. O estudo das proteases procedeu-se através de sua açäo sobre o substrato, representado pelas próprias proteínas da membrana eritrocitária. As proteases ligadas à membrana foram estudadas através de incubaçäo a 37 graus centígrados por vários períodos de tempo de estromas preparados com tampäo sem inibidor de proteases, enquanto que as proteases do citosol foram avaliadas mediante a incubaçäo a 37 graus centígrados por uma hora e 24 horas de hemolisados com estroma em suspensäo preparados com tampäo sem inibidor de proteases na presença ou näo de concentraçöes crescentes de cálcio. Após a incubaçäo e solubilizaçäo dos estromas, foi efetuada a eletroforese em gel de poliacrilamida em presença de SDS (SDS-PAGE) e posterior quantificaçäo das proteínas da membrana eritrocitária por densitometria. Os resultados obtidos permitem concluir que: 1. Existe uma protease näo dependente de cálcio, ligada à membrana do glóbulo vermelho e presente também no citosol que degrada principalmente as espectrinas, mas também a banda 2.1. Esta protease sugere tratar-se da macropaína, uma protease multicatalítica (EC 3.4.99.46). 2. Existe uma protease cálcio-dependente presente no citosol do glóbulo vermelho que degrada a banda 2.1 com 0,05 mM de cálcio e a banda 4.1 com 2,0 mM de cálcio. Esta protease deve tratar-se da calpaína I (EC 3.4.22.17), somente agindo sobre a proteína 4.1 com concentraçöes mais elevadas de cálcio que, talvez rompa ligaçöes inter-moleculares do complexo juncional passando a expor a proteína 4.1 à sua açäo. 3. A protease cálcio dependente presente no citosol näo exibiu comportamento diferente nos pacientes portadores de hipertensäo essencial, quando comparado ao grupo controle. 4. Näo houve diferenças entre os pacientes portadores de hipertensäo essencial sem tratamento e após tratamento com Captopril a Amlodipina. 5. Näo houve diferenças entre os pacientes tratados com Captopril e aqueles tratados com Amlodipina.

[Antibodies against extracellular products of group A Streptococcus. Diagnostic importance in acute rheumatic fever]. / Anticuerpos contra productos extracelulares del estreptococo grupo A. Importancia diagnóstica en la fiebre reumática aguda.

Streptococcal throat infection is a sine qua non for the development of rheumatic fever (RF) in genetically susceptible people. Demonstration of such infection is not easy. In overt RF less than 10% of patients still carry streptococci in their throat and immunologic methods are required to identify antibodies against streptococcal products (SP). Humoral response against SP was studied in children and adults with and without RF. Antistreptolysin O (ASO) showed a non-gaussian distribution, and reference value was established as percentile. Adults have a 97 percentile of 227, in children 90 percentile was 451. When RF was present all cases, except one, showed higher values. When antibodies against SP besides ASO were sought by an agglutination test (Streptozyme tm), people below 15 years of age showed low titers in 15 out of 28 cases. In contrast, high titers were the rule in children suffering RF. High ASO titer correlated with high Streptozyme value. These methods are capable to recognize an specific immune response against Group A Beta hemolytic streptococci, and are valuable tools in the diagnosis of RF.